Proteinase-Activated Receptor 4 Activation Triggers Cell Membrane Blebbing through RhoA andβ-Arrestin

Abstract

Proteinase-Activated Receptors (PARs) are a four-member family of G-protein coupled receptors that are activated via proteolysis. PAR4 is a member of this family that is cleaved and activated by serine proteinases such as thrombin, trypsin and cathepsin-G. PAR4 is expressed in a variety of tissues and cell types including platelets, vascular smooth muscle cells and neuronal cells. In studying PAR4 signalling and trafficking, we observed dynamic changes in the cell membrane with spherical membrane protrusions that resemble plasma membrane blebbing. Since non-apoptotic membrane blebbing is now recognized as an important regulator of cell migration, cancer cell invasion, and vesicular content release we sought to elucidate the signalling pathway downstream of PAR4 activation that leads to such events. Using a combination of pharmacological inhibition and CRISPR/Cas9-mediated gene-editing approaches, we establish that PAR4-dependent membrane blebbing occurs independently of the Gaq/11-and Gai-signalling pathways and is dependent on signalling via the b-arrestin-1/-2 and RhoA signalling pathways. Together these studies provide further mechanistic insight into PAR4 regulation of cellular function. Significance Statement We find that the thrombin receptor PAR4 triggers cell membrane blebbing in a RhoA-and b-arrestin-dependent manner. In addition to identifying novel cellular responses mediated by PAR4, these data provide further evidence for biased signalling in PAR4 since membrane blebbing was dependent on some, but not all, signalling pathways activated by PAR4.

Lysosomal peptidases—intriguing roles in cancer progression and neurodegeneration

“…E‐cadherin‐based cell–cell junctions are regulated by CatG promotion of E‐cadherin/catenin and E‐cadherin/protein kinase D1 complex formation and Rap1 activation in MCF‐7 cells [ 179 ]. CatG also activates proteinase‐activated receptor 4 that triggers cell membrane blebbing, a mechanism recognized as an important regulator of cell migration, cancer cell invasion, and vesicular content release [ 180 ].…”

Multiplex G Protein–Coupled Receptor Screen Reveals Reliably Acting Agonists and a Gq-Phospholipase C Coupling Mode of GPR30/GPER1

“…Since GPCR coupling to Gα 12/13 is known to affect the actin cytoskeleton through the downstream effectors RhoA and ROCK (Purvanov et al, 2014;Vanderboor et al, 2020), we applied the ROCK inhibitors Y-27632 or fasudil to affirm this assumption. Both blockers prevented bleb formation (data not shown).…”

Plasma membrane repair, blebbing and microvesiculation: parallels and relationships

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